Extracellular insulin binds to plasma membrane receptor
GLUT4 is translocated from a pre-formed intracellular pool
GLUT4-containing vesicles fuse with the plasma membrane, increasing GLUT4 on the cell surface
Defects can lead to insulin resistance and type II diabetes.
Ion Channel-Mediated
Cells must maintain electrochemical gradients to function
Gradients are a source of energy for driving many cellular processes
Ion channels are a form of facilitated diffusion
Ions can also enter by active transport via ion pumps
Transient alteration of ion membrane permeability can control cell signaling
Ion channel can pass 107 to 108 ions/second
Ion Concentration and Charge Gradients
Ion concentration varies on exterior and interior of cell
Concentrations contribute to the total charge differential across the membrane
Sum of free energy change determines direction and rate of ion flow
ΔG of ion concentration gradient plus the ΔG from electrical gradient equals the total ΔG
Most Ion channels are gated and operate in response to specific signals
i.e. voltage-gated or ligand-gated channels
Exception: K+ leak channels are always open and act to maintain the negative membrane resting potential (cytoplasm negative).
White ion channel opening responds to specific signals, the direction and flow rate relies on the relevant electrochemical gradient across the plasma membrane.
Ligand-Gated Ion Channels
Example: Nicotinic Acetylcholine Receptor – allows passage of Na+ and K+.
Nicotinic acetylcholine are at the neuromuscular junction (NMJ)
Large flux of Na+ into the cell and K+ can enter, resulting in a transient depolarization of the membrane
Leads to an action potential necessary for muscle contraction.
Uses a rotation and sliding of helices to open and close the ion channel
Voltage-Gated Ion Channels
Important for nerve impulse action potentials
Multiple related sets of α-helices will come together to form a selective ion pore.
Uses the ball-and-chain mechanism to inactivate the channel
Channel is closed when polarized
When depolarized, the channel opens
After depolarization occurs, the “ball” part of the protein inactivates the channel to stop depolarization
After repolarization, the channel closes and the “ball” returns to original conformation.
Selectivity of Ion Channels
Selectivity of ion channel depends on the structure of the channel (the amino acids in the selectivity filter)
All ions exist in a hydrated form with a distinctive water shell
Ion enters up to the point of the ion channel known as the selectivity filter.
At the selectivity filter, the channel pore size becomes too small for a hydrated ion to pass
The ion must shed its water shell in a thermodynamically favored manner, i.e. other favorable polar interactions must replace those the ion had with water.
For a K+ channel, K+ is thermodynamically favored to pass but Na+ is not.
Rate of Transport
Two-site model has two binding sites in a channel’s selectivity filter with the first ion reaching the second binding site
The next ion that binds to the first binding site and creates an electrostatic repulsion that pushes the first ion out
This process repeats to allow efficient transport of ions.
Active Transport
Require energy input to function against the electrochemical gradient.
If the transporter protein itself hydrolyses ATP, it is called primary transport
If unfavorable (uphill) flow of one molecule is coupled to a favorable (downhill) flow of another, this is called secondary transport
Primary Transport
Three families of primary active transporters: P-type, ABC-type, and F&V-type
F&V-type pumps only H+ and is common in bacteria and plants
P-Type Primary Transport
ATP-powered pumps such as Ca2+ ATPase in muscle SR, H+/K+ ATP in the stomach, Na+/K+ ATPase in all cells, among others.
Hydrolysis of ATP provides energy to pump against the electrochemical gradient
Phosphoryl group of ATP becomes covalently bound to the transporter
Requires Mg2+ cofactor
Two integral protein subunits
Example: Na+/K+ pump generates an ion concentration gradient for controlling:
Cell volume
Driving transport of sugars and amino acids
Establishing and maintaining electrochemical gradients
Maintaining membrane charge potentials at -60mV with K+ leak channels.
25%-75% of all cytosolic ATP present in cells is hydrolyzed by action of the Na+/K+
Distinct conformational states (E1 and E2) and phosphorylation state governs transport.
Cardiotonic steroid drugs (Digitalis) act by inhibiting the function of the Na+/K+ ATPase where the dephosphorylation step occurs
This locks the pump into a non-functional state, decreasing pump action and increasing Na+ in cardiac muscle cells
This leads to increase Ca2+ in the cell via action of Na+/Ca2+ transporter
Ca2+-mediated signals act to increase contraction strength of the heart muscle
ABC-Type Primary Transport
ATP-Binding Cassette Primary Transporter
Transports ions and small molecules
Has 6 α-helices form transmembrane domain and is structurally and functionally different than P-type pumps.
Functional domains (1-4) of ABC-transporters may be single or separate units.
ATP hydrolysis is coupled with solute movment
Example: MDR-1 and chemoresistant cancers
Small planar drugs diffuse into cells
Cell sees that drug as toxic and MDR-1 uses ATP to export drug from cytosol
Drugs fail to exert benefits and tumor cells become chemoresistant to multiple drugs simultaneously and lead to uncontrollable tumor growth.
MDR-1 gene is frequently amplified in multi-drug resistant cancer cells
MDR-1 inhibitors in clinical trials seems promising
MDR-1 is also found in normal tissues especially in the liver to get rid of toxins
Example: CFTR chloride channel and cystic fibrosis
Cystic fibrosis is a autosomal recessive genetic disease of mucus glands, affecting respiratory and digestive systems.
deltaF508CFTR – Deletion of 3 bp in CFTR
Mutant protein does not reach the cell surface and cannot function likely due to protein misfolding and inability for molecular chaperone to bring it to the cell surface.
Gene therapy is currently investigated to help the mutated deltaF508CFTR reach the cell surface and provide some relief to cystic fibrosis patients.
CFTR is present on apical surface of epithelial cell plasma membrane where it pumps Cl- out of the cell
deltaF508CFTR mutated cells accumulate Cl- in the cell, causing Na+ and water to come in from the extracellular space
Loss of water from extracellular space results in thick, dehydrated mucus and defective function of the respiratory tract cilia and increased infections.
Secondary Transport
Use one concentration gradient to power another gradient
Example: Na+/Glucose symport
ATP is used to pump Na+ to generate a Na+ concentration gradient
By coupling the Na+ downhill flow with the uphill of glucose, glucose can overcome its gradient and move into a cell
Antiporter – flow of A moves in the opposite direction of B
Symporter – flow of A moves in the same direction of B
Integration of Membrane Selective Permeability
Example: Glucose Transport by Intestinal Epithelia
Oral rehydration therapy depends on the function of Na+/Glucose Symporter
Both NaCl and Glucose is necessary because the symporter needs both to function
Transport of glucose and NaCl across the intestinal epithelium causes water from the intestinal lumen to move into the blood, leading to rehydration
Signal Transduction
Molecule does not actually enter a cell, just the signal such as by second messengers
Extracellular signal may be soluable (i.e. hormone or steroid) or on the plasma membrane of another cell
Endocrine – in the circulation
Paracrine – from an adjacent or nearby cell
Autocrine – from that cell itself
Stages of Signal Transduction
Signal – extracellular signal
Reception – received on the plasma membrane
Transduction – Occurs in the cell interior where receptor is phosphorylated and signaling cascades may occur
Enzymes central to signal transduction are located at the plasma membrane
Example: Adenylate Cyclase – generates cAMP from ATP
Example: Phospholipase C – generates DAG and IP3
Various signal transduction events are integrated for proper function, i.e. signal transduction has a lot of “cross-talk.”
Response – response to the signal by gene regulation, phosphorylation, etc. and can cause many levels of positive or negative feedback
Secondary Messengers
Initial signal (first messenger) is amplified and propagated by a second messenger in side the cell
Plasma Membrane II
Dr. Cynthia Smas, Ph.D.
Table of Contents
Overview
Lipid Bilayers
Cell signaling
Osmosis
Facilitated Diffusion
GLUT4 Protein
Ion Channel-Mediated
Ion Concentration and Charge Gradients
Ligand-Gated Ion Channels
Voltage-Gated Ion Channels
Selectivity of Ion Channels
Rate of Transport
Active Transport
Primary Transport
P-Type Primary Transport
ABC-Type Primary Transport
Secondary Transport
Integration of Membrane Selective Permeability
Signal Transduction
Stages of Signal Transduction
Secondary Messengers
Objectives